Purpose: To assess the alterations in conjunctival impression cytology following 23-gauge transconjunctival sutureless versus conventional pars plana vitrectomy. Patients and Methods: Sixty consecutive patients were enrolled in the study. Conjunctival impression cytology was performed on 30 eyes (of 30 subjects) with 23-gauge transconjunctival sutureless vitrectomy and on 30 eyes (of 30 subjects) with conventional vitrectomy. Conjunctival impression cytology was performed preoperatively on the 1st day and in the 3rd month after the surgery. Impression cytology specimens of each group were graded and scored using a range of 0-3 according to Nelson's method. Results: In the conventional pars plana vitrectomy group, statistically significant alterations in the conjunctival impression cytology were detected on the 1st postoperative day (p = 0.001) and in the 3rd postoperative month (p = 0.001), whereas in the 23-gauge transconjunctival sutureless pars plana vitrectomy group, statistically significant changes were observed on the 1st postoperative day. However, no significant changes were observed in the following 3 postoperative months (p = 0.08). Conclusion: The properties of impression cytology were altered in the early postoperative period after sutureless and conventional vitrectomy. These changes were improved in the sutureless vitrectomy group only. Sutureless vitrectomy also had an advantageous effect on the conjunctival cytological changes and conjunctival structure.